Human vision depends on vitamin A for light sensing by visual pigments in photoreceptor cells, the maintenance of cornea, and eye development. Insufficient uptake of vitamin A can cause severe blindness. However, vitamin A or chemicals derived from it can also be toxic if accumulated excessively. Nature devised a special delivery vehicle for vitamin A called plasma retinol binding protein (RBP), which is the principle, specific and high-affinity carrier of vitamin A in the blood. Healthy people have micromolar concentrations of RBP bound with vitamin A in the blood to satisfy the need of many biological processes that depend on vitamin A. RBP forms a one-to-one complex with vitamin A and is responsible for precise and regulated delivery of vitamin A to organs that depend on it such as the eye. The molecular machinery that absorbs vitamin A from RBP in the blood remained a mystery for more than 30 years until its recent identification as STRA6, a multi- transmembrane domain protein that has previously unknown function and is highly expressed in the retinal pigment epithelium, a cell type that plays pivotal roles in vitamin A uptake for vision. The RBP receptor represents a new type of cell-surface receptor/transporter and takes up vitamin A from RBP in the blood through a membrane transport mechanism distinct from all known mechanisms. This proposal focuses on three major unsolved questions on the molecular mechanisms of vitamin A uptake for vision. Previous studies have revealed that STRA6 has the abilities in promoting vitamin A influx into cells, efflux out of cells, or exchange between vitami A binding proteins. However, how STRA6 interacts with the transport substrate vitamin A and how STRA6's vitamin A uptake activity is regulated are unknown. Although STRA6 is the major vitamin A uptake pathway for vision, there exists an alternative pathway that takes up the ester form of vitamin A from the blood. The molecular mechanism of this vitamin A uptake pathway for vision and its relationship with STRA6-mediated vitamin A uptake are also unknown. The three specific aims in this proposal are designed to answer these questions on vitamin A uptake. All three specific aims represent major gaps in our knowledge. We have performed preliminary studies and developed several innovative techniques and reagents to make it feasible to study these specific aims.